CSMLS REVIEW HISTOLOGY EXAM
QUESTIONS WITH CORRECT
ANSWERS
Grossing - Correct Answers -described tissues (size, weight, colour)
Large tissue take section (want outer edge of bad and some normal, and lymph nodes)
small tissue do whole thing
Put in formalin to start to fix
Immunoflourescent studies not fixed
Brain fixed several days before being grossed (large container) via circle of willis
avoid cone head artifact (big enough container req)
Uterus (cut open and then fixed)
cervix is of concern and fibroids
Masectomy prolonged fixation
lung inflated with formalin
Hematology = mercury fixative (spleen)
sliced first (capsule slows fixation)
Dictation of what is in each cassette
Fixation Delay - Correct Answers -If not done properly cannot be corrected
Goal is to preserve morphology
If delayed
- cell swelling (larger), due to osmotic balance water accumulated
- glassy cytoplasm (clearer), glycogen is used up
-Eosinophilia: use of glycogen = lactic acid, lower pH, denature proteins, increase pos,
increase eosin, organelles (RER) broken down decreased Hx
- Surface changes: low pH denature filaments, lose cell attachments (Desmosomes),
desquamation (CHAOS)
,- Nuclear changes: lysosomes need exact pH to keep enzymes inactive (need E), when
gone enzymes active = nuclease
Karyolysis: Chromatin fades
Pyknosis: nucleus shrinks and dark
Karyorrhexis: Fragment nucleus
Putrefaction: bacteria proliferate, enzymes damage tissues
Autolysis: damage tissue no nuclei
Types of fixatives - Correct Answers -Chemical
- change protein shape (denature) inactive enzymes
Picric acid, formaldehyde, mercuric chloride, dichromate, osmium tetroxide, zinc salts,
alcohol
Physical
- heat removes water, changing shape inactive enzymes
Air dried (heme malignancy)
post fixed with methanol
Coating fixatives = spray covers
Fluid specimens
- centrifuge and then smear
Chemical Fixative details (coagulant) - Correct Answers -Coagulant fixatives (type of
chemical)
Alcohol (smears)
Acetic acid(w/others)inc nuclear detail
Acetone= fix frozen sections
Mercury (not used alone= compound)
enhanced nuclear staining (hematology)
toxic and corrosive
ARTIFACT black pigment (not around blood - like formalin pigment,
****remove with lugol's iodine and sodium thiosulphate)
Zinc sulfate
safer than mercury
Ag preservation, stabilizes protein conformation (histidine and cysteine)
Picric acid (phenol) (coagulation/additive)
also a stain
fix with alcohol after (keeps salt picrates that are formed)
*** explosive when dry
improved staining with anionic dyes bc acid
, has yellow colour removed with washing in high pH
Aldehyde Fixatives - Correct Answers -Non-coagulant
Formalin and glutaraldehyde
formaldehyde = gas-> saturated = aqueous formalin 100%
used at 10% formalin 3.7% formaldehyde
slow at acidic, fast in alkaline ***
paraformaldehyde = precipitate polymer white ( leads to ineffective fixation) some
methanol can prevent
Formalin pigment
Spontaneous oxidation= formic acid lower pH
in bloody tissues (when hgb breaks into acid hematein)
prevented by buffering or make fresh
** Remove with alcoholic picric acid
Formalin penetrates fast but fixes slow (24 hr (can start with formalin and finish in
alcohol to speed up)
glutaraldehyde fixation
- larger, slower penetration
- leaves free eldehyde (false PAS +)
ox quickly (keep sealed)
for electron microscopy (harder, more links = ultrastructure preserved)
fixes <2 hours
Mechanism of fixation - Correct Answers -1) formalin bind functional group and add
CH2OH (temporary, can be washed away) adding OH group = hemiacetal
2) methylene bridge formation = link functional groups via OH (24hr to form)
not easily reverse
Effects of fixation - Correct Answers -Less eosin/anionic staining
1) methylene bridges and hemiacetal involves amino groups (cationic) (less stained)
= eosin (anionic) staining reduced
2) aminos not near others = only hemiacetal no bridge
these oxidized to carboxylic acid residues (neg) increased neg decreased eosin
lipids preserved in formalin but lost in alcohols and xylene
preserves nucleic acids and nuclear proteins
No effect on carbs
Oxidizing agents fixatives - Correct Answers -Osmium tetroxide
slow
e- microscopy
QUESTIONS WITH CORRECT
ANSWERS
Grossing - Correct Answers -described tissues (size, weight, colour)
Large tissue take section (want outer edge of bad and some normal, and lymph nodes)
small tissue do whole thing
Put in formalin to start to fix
Immunoflourescent studies not fixed
Brain fixed several days before being grossed (large container) via circle of willis
avoid cone head artifact (big enough container req)
Uterus (cut open and then fixed)
cervix is of concern and fibroids
Masectomy prolonged fixation
lung inflated with formalin
Hematology = mercury fixative (spleen)
sliced first (capsule slows fixation)
Dictation of what is in each cassette
Fixation Delay - Correct Answers -If not done properly cannot be corrected
Goal is to preserve morphology
If delayed
- cell swelling (larger), due to osmotic balance water accumulated
- glassy cytoplasm (clearer), glycogen is used up
-Eosinophilia: use of glycogen = lactic acid, lower pH, denature proteins, increase pos,
increase eosin, organelles (RER) broken down decreased Hx
- Surface changes: low pH denature filaments, lose cell attachments (Desmosomes),
desquamation (CHAOS)
,- Nuclear changes: lysosomes need exact pH to keep enzymes inactive (need E), when
gone enzymes active = nuclease
Karyolysis: Chromatin fades
Pyknosis: nucleus shrinks and dark
Karyorrhexis: Fragment nucleus
Putrefaction: bacteria proliferate, enzymes damage tissues
Autolysis: damage tissue no nuclei
Types of fixatives - Correct Answers -Chemical
- change protein shape (denature) inactive enzymes
Picric acid, formaldehyde, mercuric chloride, dichromate, osmium tetroxide, zinc salts,
alcohol
Physical
- heat removes water, changing shape inactive enzymes
Air dried (heme malignancy)
post fixed with methanol
Coating fixatives = spray covers
Fluid specimens
- centrifuge and then smear
Chemical Fixative details (coagulant) - Correct Answers -Coagulant fixatives (type of
chemical)
Alcohol (smears)
Acetic acid(w/others)inc nuclear detail
Acetone= fix frozen sections
Mercury (not used alone= compound)
enhanced nuclear staining (hematology)
toxic and corrosive
ARTIFACT black pigment (not around blood - like formalin pigment,
****remove with lugol's iodine and sodium thiosulphate)
Zinc sulfate
safer than mercury
Ag preservation, stabilizes protein conformation (histidine and cysteine)
Picric acid (phenol) (coagulation/additive)
also a stain
fix with alcohol after (keeps salt picrates that are formed)
*** explosive when dry
improved staining with anionic dyes bc acid
, has yellow colour removed with washing in high pH
Aldehyde Fixatives - Correct Answers -Non-coagulant
Formalin and glutaraldehyde
formaldehyde = gas-> saturated = aqueous formalin 100%
used at 10% formalin 3.7% formaldehyde
slow at acidic, fast in alkaline ***
paraformaldehyde = precipitate polymer white ( leads to ineffective fixation) some
methanol can prevent
Formalin pigment
Spontaneous oxidation= formic acid lower pH
in bloody tissues (when hgb breaks into acid hematein)
prevented by buffering or make fresh
** Remove with alcoholic picric acid
Formalin penetrates fast but fixes slow (24 hr (can start with formalin and finish in
alcohol to speed up)
glutaraldehyde fixation
- larger, slower penetration
- leaves free eldehyde (false PAS +)
ox quickly (keep sealed)
for electron microscopy (harder, more links = ultrastructure preserved)
fixes <2 hours
Mechanism of fixation - Correct Answers -1) formalin bind functional group and add
CH2OH (temporary, can be washed away) adding OH group = hemiacetal
2) methylene bridge formation = link functional groups via OH (24hr to form)
not easily reverse
Effects of fixation - Correct Answers -Less eosin/anionic staining
1) methylene bridges and hemiacetal involves amino groups (cationic) (less stained)
= eosin (anionic) staining reduced
2) aminos not near others = only hemiacetal no bridge
these oxidized to carboxylic acid residues (neg) increased neg decreased eosin
lipids preserved in formalin but lost in alcohols and xylene
preserves nucleic acids and nuclear proteins
No effect on carbs
Oxidizing agents fixatives - Correct Answers -Osmium tetroxide
slow
e- microscopy
